Electrochemically reduced graphene oxide on CoCr biomedical alloy: Characterization, macrophage biocompatibility and hemocompatibility in rats with graphene and graphene oxide.

Electrochemically reduced graphene oxide (ErGO) films on a biomedical grade CoCr alloy have been generated and characterized in order to study their possible application for use on joint prostheses. The electrodeposition process was performed by cyclic voltammetry. The characterization of the ErGO films on CoCr alloys by XPS revealed sp2 bonding and the presence of CO and CO residual groups in the graphene network. Biocompatibility studies were performed with mouse macrophages J774A.1 cell cultures measured by the ratio between lactate dehydrogenase and mitochondrial activities. An enhancement in the biocompatibility of the CoCr with the ErGO films was obtained, a result that became more evident as exposure time increased. Macrophages on the CoCr with the ErGO were well-distributed and conserved the characteristic cell shape. In addition, vimentin expression was unaltered in comparison with the control, results that indicated an improvement in the CoCr biocompatibility with the ErGO on the material surface. The in vivo response of graphene and graphene oxide was assessed by intraperitoneal injection in wistar rats. Red blood cells are one of the primary interaction sites so hemocompatibility tests were carried out. Rats inoculated with graphene and graphene oxide showed red blood cells of smaller size with a high content in hemoglobin.

Study of overall and local electrochemical responses of oxide films grown on CoCr alloy under biological environments.

The interaction of the physiological medium and living tissues with the implant surfaces in biological environments is regulated by biopotentials that induce changes in the chemical composition, structure and thickness of the oxide film. In this work, oxide films grown on CoCr alloys at 0.5 V vs Ag/AgCl and 0.7 V vs Ag/AgCl have been characterized through overall and localized electrochemical techniques in a phosphate buffer solution and 0.3% hyaluronic acid. Nanopores of 10-50nm diameter are homogeneously distributed along the surface in the oxide film formed at 0.7 V vs Ag/AgCl. The distribution of the Constant Phase Element studied by local electrochemical impedance spectroscopy showed a three-dimensional (3D) model on the oxide films grown at 0.5 V vs Ag/AgCl and 0.7 V vs Ag/AgCl. This behaviour is especially noticeable in oxide films grown at 0.7 V vs Ag/AgCl, probably due to surface inhomogeneities, and resistive properties generated by the potentiostatic growth of the oxide film.

Analysis of metallic traces from the biodegradation of endomedullary AZ31 alloy temporary implants in rat organs after long implantation times.

AZ31 alloy has been tested as a biodegradable material in the form of endomedullary implants in female Wistar rat femurs. In order to evaluate the accumulation of potentially toxic elements from the biodegradation of the implant, magnesium (Mg), aluminium (Al), zinc (Zn), manganese (Mn) and fluorine (F) levels have been measured in different organs such as kidneys, liver, lungs, spleen and brain. Several factors that may influence accumulation have been taken into account: how long the implant has been in place, whether or not the bone is fractured, and the presence of an MgF2 protective coating on the implant. The main conclusions and the clinical relevance of the study have been that AZ31 endomedullary implants have a degradation rate of about 60% after 13 months, which is fully compatible with fracture consolidation. Neither bone fracture nor an MgF2 coating seems to influence the accumulation of trace elements in the studied organs. Aluminium is the only alloying element in this study that requires special attention. The increase in Al recovered from the sampled organs represents 3.95% of the amount contained in the AZ31 implant. Al accumulates in a statistically significant way in all the organs except the brain. All of this suggests that in long-term tests AZ31 may be a suitable material for osteosynthesis.

Fracture bone healing and biodegradation of AZ31 implant in rats.

The ideal temporary implant should offer enough mechanical support to allow healing of the fracture and then biodegrade and be resorbed by metabolic mechanisms without causing any toxic effect. The aim of this research has been to simultaneously study in situ bone healing and the biodegradation of AZ31 Mg alloy as an osteosynthesis material. The in vivo study was carried out in AZ31 implants with and without Mg-fluoride coating inserted in un-fractured and fractured femurs of Wistar rats for long experimentation time, from 1 to 13 months, by means of computed tomography, histological and histomorphometric analysis. Tomography analysis showed the bone healing and biodegradation of AZ31 implants. The fracture is healed in 100% of the animals, and AZ31 maintains its mechanical integrity throughout the healing process. Biodegradation was monitored, quantifying the evolution of gas over time by 3D composition of tomography images. In all the studied groups, gas pockets disappear with time as a result of the diffusion process through soft tissues. Histomorphometric studies reveal that after 13 months the 46.32% of AZ31 alloy has been resorbed. The resorption of the coated and uncoated AZ31 implants inserted in fractured femurs after 1, 9 and 13 months does not have statistically significant differences. There is a balance between the biodegradation of AZ31 and bone healing which allows the use of AZ31 to be proposed as an osteosynthesis material.

Modeling in vivo corrosion of AZ31 as temporary biodegradable implants. Experimental validation in rats.

In this paper, two complementary approaches, mathematical modeling and experimental results are combined to identify variables that affect the in vivo biodegradation of magnesium implants. The in vivo corrosion behavior of AZ31 alloy proposed for temporary applications as fixation of bone fractures has been modeled solving the Laplace equation by finite element method (FEM). Bar-shaped AZ31 implants of 1mm diameter and 20mm length were inserted in Wistar rat femurs with and without a fracture. The presence of gas around AZ31 implants inside the femurs has been detected in situ at the epiphysis and in fractured areas by computerized tomography (CT). Examining some in vivo conditions, the model confirms that magnesium-alloy devices have different biodegradation behavior, depending on the thickness of electrolyte at the implantation site and can be used for predicting the biodegradation behavior.

Influence of the microstructure and topography on the barrier properties of oxide scales generated on blasted Ti6Al4V surfaces.

The long-term interfacial bond between an implant and bone may be improved by creating a rough surface on the implant in order to increase the surface area available for bone/implant apposition. A natural consequence of surface roughening is an increase in metal ion release, which is itself a surface dominated process. Based on this fact, the aim of this work is to study the influence of the microstructure and topography on the barrier properties of oxide scales thermally generated at 700 °C for 1h on Ti6Al4V surfaces after blasting with Al(2)O(3) particles (coarse) or SiO(2) and ZrO(2) particles (fine). The microstructural and topographical characterization of the thermally treated blasted surfaces has been studied by means of scanning electron microscopy coupled with energy dispersive X-ray analysis, contact profilometry and X-ray diffraction. The barrier properties and corrosion behaviour of the oxide layers have been studied by means of electrochemical impedance spectroscopy (EIS) in Hank's solution. Thermal treatment at 700 °C for 1h promotes the formation of oxide scales with different morphologies and crystalline structures depending on the degree of deformation of the blasted surface. The oxide scale grown on the finely blasted sample has a pine needle-like morphology which is mainly formed of anatase TiO(2). In contrast, the oxide scale grown on the coarsely blasted sample has a globular morphology formed mainly of rutile TiO(2). The differences in morphology, i.e. fine or coarse, of the oxide scales influence the corrosion response of the blasted thermally treated samples in Hank's solution. The EIS results permit evaluation of the different oxide scales from the capacitance and resistance values obtained in the high-frequency region and show a good correlation between the morphology and barrier properties. Oxidation treatment at 700 °C for 1h of Ti6Al4V samples coarsely blasted with Al(2)O(3) improves the corrosion behaviour due to an increase in the thickness of a compact, ordered and more structurally stable oxide scale. This is due to the globular morphology of the rutile (TiO(2)) structure maintaining an average surface roughness suitable for optimal osseo-integration with long-term interfacial bonding between the implant and bone.

Corrosion behaviour of AZ31 magnesium alloy with different grain sizes in simulated biological fluids.

The corrosion behaviour of AZ31 magnesium alloy with different grain sizes immersed in simulated body fluids was compared in chloride solution (8 gl(-1)) and in phosphate-buffer solution (PBS). The influence of immersion time was also analyzed. Electrochemical techniques such as open circuit potential, polarization curves, transient currents and electrochemical impedance spectroscopy, complemented with scanning electron microscopy and energy dispersive spectroscopy, were used. Immediately after the immersion in the corrosive media the corrosion resistance was similar for both grain sizes of the AZ31 alloy and higher in NaCl solutions than in PBS. However, this corrosion behaviour was reversed after longer periods of immersion due to the stabilizing of the corrosion products of MgO by P-containing compounds. These P-compounds contribute to a higher level of protection by hindering the aggressive action of chloride ions. The best corrosion behaviour of the AZ31 alloy was obtained for the finest grain alloy associated with the highest transfer resistance value, after long periods of immersion in PBS.

Study of the interaction of inorganic and organic compounds of cell culture medium with a Ti surface.

The interaction between Ti and each component of Dulbecco's modified Eagle's medium was studied in depth using different techniques, such as the measurement of the corrosion potential, electrochemical impedance spectroscopy and polarization curves. The characterization of metal surfaces was carried out by scanning electron microscopy and X-ray photoelectron spectroscopy (XPS). The adsorption process of each component was studied using the quartz crystal balance (QCM). The QCM and XPS results reveal that the adsorption kinetics for phosphate and calcium ions is slow. However, the bovine serum albumin (BSA) totally covers the Ti surface rapidly. Because the passive film (titanium oxide) has acidic hydroxyl groups, the calcium ions would have a bridging effect on the electrostatic adsorption of phosphate ions as well as that of BSA. The polarization curves reveal that the adsorbed glucose permits the ionic diffusion of the oxygen to the electrode, while the BSA and fetal bovine serum (FBS) adsorbed after 7 days of immersion act as a diffusive barrier. The impedance measurement and data fitting to the electrical equivalent circuit model show that the resistance of the proteins/TiO(2) interface, for Ti immersed in FBS, is higher than those obtained for BSA, due to the proteins present in the solution as well as the fact that the adsorbed proteins on the surface are greater.

In situ cell culture monitoring on a Ti-6Al-4V surface by electrochemical techniques.

In this work, the in situ interaction between Ti-6Al-4V alloy and osteoblastic cells has been studied by electrochemical techniques as a function of time. The interaction has been monitored for cell adhesion and growth of human osteoblastic Saos-2 cells on Ti-6Al-4V samples. The study has been carried out by electrochemical techniques, e.g., studying the evolution of corrosion potential with exposure time and by electrochemical impedance spectroscopy. The impedance results have been analyzed by using different equivalent circuit models that simulate the interface state at each testing time. The adhesion of the osteoblastic cells on the Ti-6Al-4V alloy leads to surface areas with different cell coverage rates, thus showing the different responses in the impedance diagrams with time. The effect of the cells on the electrochemical response of Ti-6Al-4V alloy is clearly seen after 4 days of testing, in which two isolated and well-differentiated time constants are clearly observed. One of these is associated with the presence of the cells and the other with a passive film on the Ti-6Al-4V alloy. After 7 days of culture, the system is governed by a resistive component over a wide frequency range which is associated with an increase in the cell coverage rate on the surface due to the extracellular matrix.

Concentration-dependent effects of titanium and aluminium ions released from thermally oxidized Ti6Al4V alloy on human osteoblasts.

Thermal oxidation treatments of Ti6Al4V, at 500 and 700 degrees C, for 1 h result in the formation of an outer "ceramic" layer of rutile, which enhances osteoblast response. In the present study, we have measured in vitro Ti and Al ion release from Ti64 alloy in the as-received state and after thermal oxidation treatments at 500 or 700 degrees C, to culture medium under standard cell-culture conditions. Concentrations of both Ti and Al released from both thermal oxidation treatments were lower than from polished alloy. Al was released from the treated or untreated surfaces in substantially lower extent than Ti. Titanium and aluminium ions affected primary human osteoblast proliferation, metabolic activity, and differentiation in a dose-dependent manner. Treatments with individual Ti or Al metal ions in similar concentration ranges than released from the surfaces did not alter osteoblast response, which also remained unaffected after treatments with combinations of Ti plus Al applied in the proportional relations than detected in ion-release experiments. We then selected higher concentrations of Ti that impaired osteoblast proliferation and differentiation, while the proportional lower concentrations of Al did not alter osteoblast behavior. In spite of its inert character, it was found that Al significantly enhanced the deleterious effect of Ti on osteoblast differentiation. Therefore, thermal oxidation treatments of Ti6Al4V alloy may improve the biocompatibility of the alloy by reducing both Ti and Al release, and thus attenuating ion-mediated interference with osteoblast differentiation.

In vitro corrosion behaviour and osteoblast response of thermally oxidised Ti6Al4V alloy.

In this work, the influence of thermal oxidation treatments of Ti6Al4V at 500 degrees C and 700 degrees C for 1 h on the in vitro corrosion behaviour and osteoblast response is studied. The potential of these treatments, aimed to improve the wear surface performance as biomaterial, relies in the formation of an outer "ceramic" layer of rutile. The corrosion behaviour was evaluated in simulated human fluids by electrochemical impedance spectroscopy and anodic polarisation tests. The effect of these thermal oxidation treatments on osteoblastic behaviour was studied in primary cultures of human osteoblastic cells. Results show that thermal oxidation treatments do not decrease the high in vitro corrosion resistance of the Ti6Al4V alloy. Osteoblast adhesion studies indicate that thermal oxidation treatments do not impair the material biocompatibility. Moreover, the thermal oxidation at 700 degrees C enhances the in vitro osteoblastic cell attachment compared to the thermal oxidation at 500 degrees C.

A surface modified ODS superalloy by thermal oxidation for potential implant applications.

In the present work attention is paid on the composition, structure and protective properties of alumina layer produced by high temperature oxidation on MA 956 superalloy (Fe-20Cr-4.5Al-0.5Ti-0.5Y(2)O(3) (wt %)). The combination of good mechanical properties of this material and the excellent biocompatibility, the good wear and corrosion behavior of an outer alpha-alumina layer, limiting the release of ionic species and wear debris from the bulk material into the body-fluid environment, can make this material a candidate alloy for medical applications. Isothermal oxidation at 1100 degrees C in air of the alloy has led to the formation of a fine-grained, compact and adherent alpha-alumina scale. Oxide nodules rich in Ti, Y, Cr, and Fe were found on the top of the surface. In vitro electrochemical corrosion experiments showed good protective properties of the oxide scale. Moreover, no spallation of the alumina layer was observed. This feature is significant considering that the alumina layer has to withstand very high compressive stresses resulting from both growth and thermal stresses incorporated during cooling.

Effect of substrate roughness on the corrosion behaviour of the Al2O3/MA 956 system.

This paper presents the influence of substrate roughness on the corrosion behaviour of the Al2O3/MA 956 system. An alumina layer of thickness 1-5 microm was generated of the MA956 alloy by thermal oxidation at 1100 degrees C using different exposure times. This Al2O3/MA 956 system with a polished substrate has shown excellent corrosion behaviour in a physiological fluid, due to the fact that the alpha-Al2O3 layer formed is dense, continuous and firmly adhered to the substrate, irrespective of the scale thickness. This good adherence allows it to withstand potentials above 1.7 V. Specimens with rough finish substrate and treatment times above 10 h present spallation of the alumina layer at the crests of the roughness profile. In this case a mixed corrosion behaviour between an alumina coated material and one with a passive layer is observed. In both types of specimens, rough and smooth, once the passivation layer is broken the repassivation capacity of the substrate is ensured due to the high chromium content of the alloy, under oxygenation conditions.

Optimal conditions for alumina coating formation on the MA956 superalloy for prosthetic bearing applications.

An experimental study of the oxidation treatment at high temperature of the ODS MA956 superalloy was conducted in an attempt to achieve a protective alumina scale for biomedical applications. A quadratic response-surface model was developed in order to study the effects of treatment time and temperature (in the range of 1000 degrees C to 1250 degrees C) on scale thickness. The obtained model adequately represents the experimental response and shows that the thickness gradients of the layer increase with the temperature for each exposure time and decrease steadily to zero as the treatment time increases. The microstructural characterization reveals that the alumina scale formed at or above 1000 degrees C consists of an alpha-alumina phase. Treatments at temperatures above 1150 degrees C give rise to an alumina scale with some defect probability. An increase in the temperature up to 1200 degrees C gives rise to the appearance of some blistering of the superficial scale. An oxidation treatment of 100 h at 1100 degrees C was found to be the best for guaranteeing the formation of a defect-free, compact, adherent, and continuous alpha-alumina scale thick enough to support satisfactory wear and biological conditions.

Comparative study of the corrosion behavior of MA-956 and conventional metallic biomaterials.

In this work the corrosion behavior of a new biomaterial, the MA-956 superalloy, immersed in Hank's solution is evaluated. A comparison with conventional metallic alloys used as articular implants is established. To determine the corrosion behavior we employed electrochemical methods: evaluation of corrosion potential Ecorr, electrochemical impedance spectroscopy (EIS), and anodic polarization curves. The corrosion resistance of the MA-956 superalloy preoxidized at 1100 degrees C during 100 h is at least two orders of magnitude higher than for the other alloys. This satisfactory behavior is stationary with time. Also the probability of the appearance of the pitting corrosion process is very low. When cracking is generated in the alpha-alumina layer the repassivation process is assured because of the high Cr content in the superalloy. This study is the first step in proposing this new alloy as a biomaterial. The low toxicity of these metallic alloys in the physiological environment suggests that in vivo their biocompatibility could be satisfactory.